Polymorphonuclear leukocytes (PMNs) and macrophages require a contractile apparatus for shape changes, movement, and phagocytosis. The major component of this motor is actin. Actin can exist as monomers or as filaments of varying lengths. Changes in actin filament concentration and length underlie the ability of these cells to move. The present studies should begin to clarify how phagocytes control actin filament assembly in their cytoplasm. (1) Studies of acumentin will be continued. This 65,000 dalton protein purified from macrophages and human granulocytes controls actin assembly by blocking actin monomer exchange at the low affinity end of actin filaments. Conditions which enhance and inhibit this activity will be assessed by viscometry. Acumentin's binding affinity for actin will be measured using a radiolabel sedimentation assay. Its presence in other cells and tissues will be demonstrated by nitrocellulose immunoblot. Acumentin's localization in phagocytes will be visualized by immunofluorescence (2) Actin assembly in various combinations of macrophage actin-modulating proteins (acumentin, profilin, gelsolin and any newly purifed proteins) will be studied by light acattering, pyrenyl-actin, DNAse I inhibition, and viscosity. (3) To see if recombination studies mimic whole extracts, PMN and macrophage extracts will be studied using a new fluorescent probe, pyrenyl-actin. Preliminary studies suggest the presence of a new calcium sensitive actin-modulating protein whose function is different than gelsolin's. (4) This new calcium sensitive regulatory protein will be purified by ion exchange and gel filtration chromatography. (5) A granulocyte actin polymerization defect discovered in two male children will be studied. These children also had a severe ultimately fatal PMN chemotactic defect. Actin will be purified from the granulocytes of one child's parents and subjected to thin layer and high pressure liquid chromatography peptide mapping. Understanding how actin works in phagocytes may lead to new methods for altering leukocyte movement and thereby improve our ability to enhance host defence and control inflammation.